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Abstract

Objective—It is unknown whether elevated maternal low-density lipoprotein cholesterol (LDL-C) levels lead to dyslipidemia in the offspring. Because this could have important consequences for cardiovascular prevention in mother and child, we explored the relationship between maternal familial hypercholesterolemia (FH) and lipids in adult offspring.

Methods and Results—In a large cohort of both Dutch and Canadian origin, we compared lipid profiles between patients, aged 18 to 85 years, who inherited FH maternally (n=1069) and those who inherited FH paternally (n=1270). This relationship was evaluated using multivariate regression analyses. Levels of total cholesterol (TC), LDL-C, and apolipoprotein B 100 (ApoB100) were significantly elevated in patients who inherited FH maternally compared with patients who inherited FH paternally (adjusted differences in TC: 0.156 mmol/L, P=0.037; LDL-C: 0.187 mmol/L, P=0.012; ApoB: 0.064 g/L, P=0.022).

Conclusion—Our data show that maternal hereditary hypercholesterolemia slightly increases TC, LDL-C, and ApoB levels in their offspring later in life. Although the molecular mechanisms underlying these observations still require elucidation, our data suggest that maternal hypercholesterolemia during pregnancy may program lipid metabolism to a certain extent in the fetus.

Familial hypercholesterolemia (FH) is a common autosomal dominant disorder of lipoprotein metabolism, caused by mutations in the low-density lipoprotein receptor (LDLR) gene.1 As a result, FH patients are exposed to elevated levels of low-density lipoprotein cholesterol (LDL-C) from birth onward and are subsequently predisposed to premature cardiovascular disease (CVD). Lowering LDL-C levels in these patients by means of lipid lowering treatment can significantly prevent or delay the onset of CVD and premature death.2

In women with FH who wish to become pregnant, the current advice is to discontinue statin therapy to avoid teratogenesis in the unborn infant and to reinstitute statin therapy after lactation is finished.3,4 Consequently, serum cholesterol levels exhibit a sharp increase, and during gestation this becomes even more pronounced.5 Several studies suggest that maternal cholesterol is transported from the maternal to the fetal circulation6,7 and that lipid levels of the mother are closely aligned to those of the fetus in the first 6 months of pregnancy.8 As a result, fetuses of FH mothers may be exposed to rather high lipid levels.

Notably, animal and human studies suggest that there may be a critical window for fetal development where changes in the maternal condition can influence long-term cardiovascular risk in the offspring. At one end of the spectrum, maternal nutrient deprivation is associated with a more atherogenic lipid profile and increased cardiovascular risk in the offspring in adult life.9,10 Conversely, maternal hyperglycemia, for instance, has been associated with a higher risk of developing metabolic syndrome in the offspring.11 There are also indications that, besides hyperglycemia, maternal hypercholesterolemia may increase the risk for CVD in the offspring.12–15 A recent study showed that adult mice born to hypercholesterolemic dams had an increased transcriptional activity of both genes involved in the cholesterol synthesis and LDLR in the liver.15 In humans, fatty streak formation progressed strikingly faster in children of hypercholesterolemic mothers than in those of normocholesterolaemic mothers,16 but the long-term consequences in humans are still controversial.16–18

From a public health and prevention perspective, it would be important to establish whether maternal hypercholesterolemia can influence lipid levels and subsequent CVD risk in the offspring. The FH population would constitute an excellent model to evaluate this possible association. To do so, we collected data on the FH carrier status of the parents in a large group of FH patients and explored the association between maternal FH and lipid profiles in their offspring.

Methods

Study Population

For the current study, we used data on FH patients of both Dutch and French Canadian origin. The Dutch patients participated in the Genetic Identification of Risk Factors in FH study.19 This was a retrospective multicenter cohort study designed to determine the influence of genetic factors on disease progression of FH. Four thousand hypercholesterolemic patients were randomly selected from the central DNA and Biobank at the Academic Medical Centre Amsterdam, the Netherlands. Patients who were older than 18 years and fulfilled the criteria for FH were selected for the cohort. Furthermore, only 1 index case per family was included. Altogether, 2400 FH patients from 27 outpatient lipid clinics made up this cohort.

The French Canadian group consisted of 1037 consecutive FH patients aged 18 years and older who visited the Chicoutimi Hospital Lipid Clinic between 1993 and 2005.

The diagnosis FH in both countries was based on similar criteria: (1) presence of a documented LDLR mutation, or (2) plasma LDL-C levels above the 95th percentile for age and gender in combination with at least 1 of the following: the presence of typical tendon xanthomata in the patient or in a first-degree relative, an LDL-C level above the 95th percentile for age and gender in a first-degree relative, or proven coronary artery disease in the patient or in a first-degree relative under the age of 60 years.

Patients in which the mode of inheritance of FH was unknown were excluded for the current study, as were those for whom no (untreated) lipid data were available above the age of 18 years and those whose triglycerides (TG) were above 5 mmol/L.

Information about demographic characteristics, classical risk factors, and lipids for the Dutch patients was collected from the patient’s medical records, and smoking habits were collected from supplemental questionnaires sent to the patient.19 For the French Canadian patients, these data were obtained from patients at their first visit at the Chicoutimi Hospital Lipid Clinic. Informed consent was obtained from all patients.

Laboratory Parameters

Lipid and lipoprotein levels were determined in fasting patients not using lipid lowering medication for at least 6 weeks in the Dutch and 2 to 4 weeks in the French Canadian patients. Plasma total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), and TG concentrations were measured by standard methods, and LDL-C concentrations were calculated using the Friedewald formula. Plasma apolipoprotein B100 (ApoB) and apolipoprotein A1 (ApoA1) concentrations were measured by standard methods in the French Canadian patients; these data were not available for the Dutch patients. Apolipoprotein E (ApoE) genotypes were identified by characteristic visible bands after amplification by polymerase chain reaction, restriction endonuclease digestion, and electrophoresis on 5% agarose gel.20

Statistical Analyses

The Dutch and French Canadian patients were all patients from outpatient lipid clinics and fulfilled the same inclusion criteria. Therefore, we decided to combine these patients for the statistical analyses. Differences in variables with a continuous or a dichotomous distribution between patients who inherited FH maternally and those who inherited FH paternally were evaluated using linear or logistic regression analyses, respectively. In a stepwise backward multivariate regression analysis, we considered the following potential confounders: gender, age, body mass index, smoking, diabetes mellitus, ApoE genotype, parental history of CVD, country, and lipids. The analyses were performed using the generalized estimating equation method in the SAS procedure GENMOD to account for correlations within families. The exchangeable correlation structure was used for these models. Variables with a skewed distribution were log-transformed before statistical analyses. Probability values <0.05 were considered to indicate statistical significance. The analyses were performed with the SAS package version 9.1 (SAS Institute Inc, Cary, NC).

Results

General Characteristics

In 895 patients, the mode of inheritance was not known, and in 203 patients, baseline (untreated) lipid levels were not available. These patients were excluded for the current analyses. In total, 2339 FH patients aged between 18 and 85 years were eligible, of whom 1069 patients had inherited FH maternally and 1270 paternally. The general characteristics did not differ between these 2 groups, except for male gender (46% in patients who inherited FH maternally versus 51% in those who inherited FH paternally; P=0.006) and paternal history of CVD (55% versus 75%, respectively; P<0.001) (Table 1). Furthermore, ApoE genotype (E3E4/E4E4) showed a trend toward a statistically significant difference, ie, 31% versus 27%, respectively; P=0.062 (more detailed information on ApoE genotype can be found in supplemental Table I, available online at http://atvb.ahajournals.org). The mean number of relatives was similar for patients who inherited FH maternally and those who inherited FH paternally (1.28±0.8 and 1.27±0.8; P=0.706).

Discussion

This study shows that adult offspring of FH mothers had slightly but statistically significant increased levels of TC, LDL-C, and ApoB compared with offspring of FH fathers. These findings indicate that patients who inherit FH through their mother may have a more atherogenic lipid profile than those who inherit FH from their father.

Our data suggest that maternal hypercholesterolemia similarly affects LDL-C levels in the offspring later in life as the contrary condition of poor maternal nutrition and consequential fetal undernutrition does.9 However, our findings are not unexpected. In a study with ApoE-deficient mice crossbred with wild-type mice, the heterozygous offspring born to ApoE-deficient dams had an almost 3-fold increase in TC levels compared with heterozygous mice born to wild-type dams at the age of 8 months.21 Moreover, plaque formation was increased in offspring born to hypercholesterolemic dams.14,21 In a human study, the effects of exposure to diabetes in utero were evaluated in the macrosomic offspring of both diabetic and healthy mothers. The diabetic mothers had significantly higher total and LDL-C levels (219.8 versus 146.5 mg/dL) compared with the healthy mothers, and their macrosomic offspring had significantly higher LDL-C levels than macrosomic neonates of healthy mothers (126.1 versus 74.2 mg/dL).22 In accordance with this, increased maternal LDL-C levels were associated with an upregulation of cholesterol synthesis in the offspring.18 This may subsequently cause a more atherogenic lipid profile and a higher susceptibility for atherosclerosis later in life. Indeed, Napoli et al showed that atherogenesis was more pronounced in children or adolescents of hypercholesterolemic mothers.16 Altogether, we speculate that cholesterol synthesis and excretion is to some extent affected by maternal cholesterol levels during pregnancy in utero, which may cause increased LDL-C levels and, subsequently, an increased risk for the development of atherosclerosis in the offspring later in life.

Our findings are in contrast with results of previous human studies. Napoli et al showed that lipid levels in children from hypercholesterolemic mothers do not differ from those of children from normocholesterolemic mothers.16 Tonstad et al found no statistically significant differences in LDL-C levels between the children who inherited FH maternally or paternally (6.6 and 6.5 mmol/L, respectively).23 In addition, some other studies found that lipid levels in cord blood of newborns are not affected by the maternal cholesterol profile.17,18,24 For instance, Descamps et al showed that lipoprotein concentrations in cord blood of newborns depend primarily on the genetic status of the mothers and are related neither to the inheritance of the maternal polymorphisms by the newborns nor to the maternal lipoprotein changes caused by these polymorphisms.24 An explanation for the conflicting effects of maternal hypercholesterolemia on lipid levels in the offspring between other studies and ours may lie in the study design, size, and population. All the abovementioned studies included small numbers of (FH) subjects and focused on the effect of maternal hypercholesterolemia in newborns or children only. Our study, however, focused on the long-term effect of maternal hypercholesterolemia, because we included only adults who, in addition, all had FH themselves.

The exact mechanisms underlying the effects of maternal hypercholesterolemia in the offspring are still unclear. However, there is increasing evidence that epigenetic programming of metabolism during embryonal or fetal development might be involved.25 Epigenetic phenomena occur at the interface between the genome and the environment. The environment can influence epigenetic information that is superimposed on the DNA, which may have long-term consequences for the transcription of specific regions of the genome. Results of animal studies show that permanent changes in either DNA methylation or chromatin modification status or both may be responsible for the epigenetic programming of increased atherosclerotic susceptibility.25 For instance, maternal hypercholesterolemia in ApoE-deficient mice leads to activation of genes involved in cholesterol synthesis, as well as LDLR, in the adult offspring.15 Other animal studies have shown that the genes involved in immune pathways and fatty acid metabolism are upregulated in the offspring from hypercholesterolemic dams.14,25 These findings indicate that an adverse maternal environment is programmed in basic cellular processes of the fetus.25 Further research is needed to unravel the exact mechanisms by which maternal hypercholesterolemia influences epigenetic fetal programming.

Some methodological aspects of our study merit discussion. First of all, our data are a result of post hoc analyses. Our cohort was not designed to study our hypothesis, and consequently, data collection with respect to details on the mode of inheritance might have received less attention. In 1/3 of the patients, data about mode of inheritance and lipids were missing, leading to elimination of these individuals from the analysis. However, the excluded patients showed similar baseline characteristics compared with the included FH patients (data not shown), which makes selection bias unlikely. Furthermore, we adjusted for confounders by means of multiple regression analyses where appropriate, but some of the potential confounders lacked data. For example, there were more females among the subjects who inherited FH maternally, which might have confounded the association between lipids and FH inheritance because plasma lipid levels also depend on the menopausal status of the women.26 We knew menopausal status of 95% of the French Canadian females, but we had no such information of the Dutch females. Therefore, in case menopausal status was missing, we estimated the percentage postmenopausal or menopausal patients by assuming that all female patients above 50 were postmenopausal or menopausal. The proportions of (post) menopausal females did not differ between the patients who inherited FH maternally and those who inherited FH paternally (19% versus 18%, respectively; P=0.5). Furthermore, when we adjusted for this variable in a stepwise multiple linear regression analyses to evaluate the association between lipid levels and the mode of inheritance, (post)menopausal status did not appear to be a significant confounder. In addition, another potential confounder that lacked data were the type of LDLR mutation, which is known to affect plasma lipid profile. In the patients with a known LDLR mutation, the various mutations were equally divided between those who inherited FH paternally and those who inherited FH maternally (supplemental Table II). We also divided the mutations in receptor-negative and receptor-defective mutations, and these were also equally divided between the 2 groups (31% versus 32%, respectively; P=0.4). However, because of the extent of missing information, we feel that it is not possible to draw valid conclusions on the potential confounding effect of the type of LDLR mutation on the association between plasma lipid profile and FH inheritance. However, we are convinced that this should definitely be explored in future research. Another point is that the prevalence of parental history of CVD strikingly differed between the patients who inherited FH from their mother and patients who inherited FH from their father (Table 1). This was highest in the group of FH patients born from FH fathers, which can be explained by the tendency of (FH) women to get heart disease later in life than (FH) men.27 However, by means of multiple regression analyses, parental history of CVD appeared not to be an independent predictor of lipids, and this implies that parental history of CVD did not affect our findings. Finally, we realize that the differences in LDL-C and ApoB levels between the patients who inherited FH maternally and those who inherited it paternally are rather small; however, all differences pointed toward the same direction. Notably, differences are on the same order of magnitude as shown by Roseboom et al, who showed that the contrary condition, ie, poor maternal nutrition and consequential fetal undernutrition during the Dutch famine, results in a more atherogenic lipid profile.10 In our opinion, the differences found in this FH population therefore should not be neglected. Nevertheless, we realize that these findings should be interpreted with caution and that their clinical relevance needs further confirmation. We therefore aim to repeat our analyses in other cohorts.

Our study cohort comprised patients of Dutch and Canadian origin. Although there were differences between the two countries (eg, in mutations), these were equally distributed among patients born from FH mothers and FH fathers (supplemental Table III). Moreover, the included participants were all a random sample of patients from an outpatient lipid clinic and fulfilled the same criteria for the diagnosis of FH. Therefore, we decided to combine the 2 populations before we performed the statistical analyses. To test the robustness of our results, we also performed the analyses separately for the Dutch and the Canadian cohorts, which led to similar conclusions (data not shown).

In conclusion, our data show that maternal hypercholesterolemia due to FH is associated with slightly higher TC, LDL-C, and ApoB levels in their adult offspring. Although the differences are rather small, they all point in the same direction, indicating that maternal FH leads to a more atherogenic lipid profile, which may increase the already existing CVD risk in the FH offspring. Although the molecular mechanisms underlying these observations still require elucidation, our data suggest that maternal hypercholesterolemia during pregnancy may program lipid metabolism to a certain extent in the fetus.

Acknowledgments

We thank Dr H.R. Büller and Dr H. Scheffer for the careful reading of our manuscript and their valuable suggestions.

Sources of Funding

The Dutch Genetic Identification of Risk Factors in FH study was supported by a grant from the Netherlands Heart Foundation (98/165). The recruitment and phenotyping of the French Canadian patients was supported by ECOGENE-21 (Canadian Institutes of Health Research Grant CTP-82941) and AstraZeneca Canada Inc.